左旋多巴对c17.2神经干细胞的毒性作用及培高利特的保护作用

Cytotoxicity of Levodopa on c17.2 neural stem cell and neuroprotection of Pergolide

目的探讨左旋多巴对c17.2神经干细胞的毒性作用和培高利特的保护作用。方法c17.2神经干细胞加左旋多巴和培高利特培养不同时间,观察各组细胞的存活率、凋亡率和p53、Bax、Bcl-2、核因子(NF)-κBp65、细胞色素C、半胱氨酸蛋白酶(caspase-3)和caspase-3活性片段的表达。结果左旋多巴可使c17.2神经干细胞活性降低,与剂量相关(均P<0.05),Brdu标记显示200μmol/L左旋多巴可使细胞增殖活力下降,与时间相关(P<0.05),作用12h、24h后的Annexin-V染色荧光显微镜和流式细胞仪可检测到凋亡细胞,Western Blot显示p53、Bax、细胞色素C、caspase-3和caspase-3活性片段表达量增加。培高利特能部分保护神经干细胞免受左旋多巴的影响(均P<0.05),并能部分抑制左旋多巴引起的细胞凋亡(均P<0.05)。结论大剂量左旋多巴对c17.2神经干细胞具有毒性作用,呈剂量和时间依赖性。左旋多巴通过抑制DNA合成影响细胞增殖,并通过活化p53、Bax、细胞色素C、caspases-3途径促进细胞凋亡,培高利特可部分拮抗左旋多巴的毒性作用。

Objective To investigate the toxicity of Levodopa on c17.2 neural stem cell and neuroprotection of pergofide. Methods c17.2 neural stem cells were cultured with Levodopa and pergolide for different time point, cell survival and apoptosis rates and expressions of p53, Bax, Bcl-2, nuclear factor （NF）-KB p65, Cytochrome C, caspase-3 and the cleavage of caspase-3 were measured. Results Levodopa induced a concentration- and timedependent decrease of viability and proliferation in c17. 2 neural stem cells （ all P 〈 O. 05 ）. There were some apoptotic cells dyed by Annexin-V could be found under fluorescent microscope and flow cytometry at 12 and 24 hours following exposure to Levedopa 200 μmol/L. The elevated p53, Bax, Cytochrome C, caspase-3 and the cleavage of easpase-3 were demonstrated in c17.2 neural stem cells exposed to Levodopa. These alterations could be partly inhibited by Pergolide （ all P 〈 0. 05 ）. Condusions The concentrations of 1〉 200 μmol/L Levodopa are neurotoxic to el7.2 neural stem cells through inhibition of DNA synthesis and cell proliferation. The activation of p53, Bax, Cytoehrome C and easpases-3 protease may contribute to the mechanism by which Levodopa induces cell apoptosis. Pergolide, an anti-Parkinson drug, may partly block Levodopa-induced cytotoxicity as a neuroprotective effect.